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Article: Topological analysis of a haloacid permease of a Burkholderia sp. bacterium with a PhoA-LacZ reporter

TitleTopological analysis of a haloacid permease of a Burkholderia sp. bacterium with a PhoA-LacZ reporter
Authors
Issue Date2009
PublisherBioMed Central Ltd. The Journal's web site is located at http://www.biomedcentral.com/bmcmicrobiol/
Citation
Bmc Microbiology, 2009, v. 9 How to Cite?
AbstractBackground: 2-Haloacids can be found in the natural environment as degradative products of natural and synthetic halogenated compounds. They can also be generated by disinfection of water and have been shown to be mutagenic and to inhibit glyceraldehyde-3-phosphate dehydrogenase activity. We have recently identified a novel haloacid permease Deh4p from a bromoacetate- degrading bacterium Burkholderia sp. MBA4. Comparative analyses suggested that Deh4p is a member of the Major Facilitator Superfamily (MFS), which includes thousands of membrane transporter proteins. Members of the MFS usually possess twelve putative transmembrane segments (TMS). Deh4p was predicted to have twelve TMS. In this study we characterized the topology of Deh4p with a PhoA-LacZ dual reporters system. Results: Thirty-six Deh4p-reporter recombinants were constructed and expressed in E. coli. Both PhoA and LacZ activities were determined in these cells. Strength indices were calculated to determine the locations of the reporters. The results mainly agree with the predicted model. However, two of the TMS were not verified. This lack of confirmation of the TMS, using a reporter, has been reported previously. Further comparative analysis of Deh4p has assigned it to the Metabolite:H+ Symporter (MHS) 2.A.1.6 family with twelve TMS. Deh4p exhibits many common features of the MHS family proteins. Deh4p is apparently a member of the MFS but with some atypical features. Conclusion: The PhoA-LacZ reporter system is convenient for analysis of the topology of membrane proteins. However, due to the limitation of the biological system, verification of some of the TMS of the protein was not successful. The present study also makes use of bioinformatic analysis to verify that the haloacid permease Deh4p of Burkholderia sp. MBA4 is a MFS protein but with atypical features. © 2009 Tse et al; licensee BioMed Central Ltd.
Persistent Identifierhttp://hdl.handle.net/10722/89310
ISSN
2015 Impact Factor: 2.581
2015 SCImago Journal Rankings: 1.391
PubMed Central ID
ISI Accession Number ID
Funding AgencyGrant Number
University Seed Funding Programme for Basic Research 2008
Research Grants Council of the Hong Kong Special Administrative Region, ChinaHKU7536/06 M
Funding Information:

We thank Herbert Winkler for plasmid pMA632 and Janice Brabyn for reading the manuscript. YMT thanks the University of Hong Kong for a studentship. We gratefully acknowledge the support of the BIOSUPPORT project http://bioinfo.hku.hk for providing bioinformatics resources and computational services from the HKU Computer Centre. This work was supported by the University Seed Funding Programme for Basic Research 2008 and the Research Grants Council of the Hong Kong Special Administrative Region, China (project no. HKU7536/06 M).

References

 

DC FieldValueLanguage
dc.contributor.authorTse, YMen_HK
dc.contributor.authorYu, Men_HK
dc.contributor.authorTsang, JSHen_HK
dc.date.accessioned2010-09-06T09:55:14Z-
dc.date.available2010-09-06T09:55:14Z-
dc.date.issued2009en_HK
dc.identifier.citationBmc Microbiology, 2009, v. 9en_HK
dc.identifier.issn1471-2180en_HK
dc.identifier.urihttp://hdl.handle.net/10722/89310-
dc.description.abstractBackground: 2-Haloacids can be found in the natural environment as degradative products of natural and synthetic halogenated compounds. They can also be generated by disinfection of water and have been shown to be mutagenic and to inhibit glyceraldehyde-3-phosphate dehydrogenase activity. We have recently identified a novel haloacid permease Deh4p from a bromoacetate- degrading bacterium Burkholderia sp. MBA4. Comparative analyses suggested that Deh4p is a member of the Major Facilitator Superfamily (MFS), which includes thousands of membrane transporter proteins. Members of the MFS usually possess twelve putative transmembrane segments (TMS). Deh4p was predicted to have twelve TMS. In this study we characterized the topology of Deh4p with a PhoA-LacZ dual reporters system. Results: Thirty-six Deh4p-reporter recombinants were constructed and expressed in E. coli. Both PhoA and LacZ activities were determined in these cells. Strength indices were calculated to determine the locations of the reporters. The results mainly agree with the predicted model. However, two of the TMS were not verified. This lack of confirmation of the TMS, using a reporter, has been reported previously. Further comparative analysis of Deh4p has assigned it to the Metabolite:H+ Symporter (MHS) 2.A.1.6 family with twelve TMS. Deh4p exhibits many common features of the MHS family proteins. Deh4p is apparently a member of the MFS but with some atypical features. Conclusion: The PhoA-LacZ reporter system is convenient for analysis of the topology of membrane proteins. However, due to the limitation of the biological system, verification of some of the TMS of the protein was not successful. The present study also makes use of bioinformatic analysis to verify that the haloacid permease Deh4p of Burkholderia sp. MBA4 is a MFS protein but with atypical features. © 2009 Tse et al; licensee BioMed Central Ltd.en_HK
dc.languageengen_HK
dc.publisherBioMed Central Ltd. The Journal's web site is located at http://www.biomedcentral.com/bmcmicrobiol/en_HK
dc.relation.ispartofBMC Microbiologyen_HK
dc.subject.meshAlkaline Phosphatase - genetics-
dc.subject.meshAmino Acid Sequence-
dc.subject.meshBacterial Proteins - genetics-
dc.subject.meshBurkholderia - enzymology - genetics-
dc.titleTopological analysis of a haloacid permease of a Burkholderia sp. bacterium with a PhoA-LacZ reporteren_HK
dc.typeArticleen_HK
dc.identifier.openurlhttp://library.hku.hk:4550/resserv?sid=HKU:IR&issn=1471-2180&volume=9 article no. 233&spage=&epage=&date=2009&atitle=Topological+analysis+of+a+haloacid+permease+of+a+Burkholderia+sp.+bacterium+with+a+PhoA-LacZ+reporteren_HK
dc.identifier.emailTsang, JSH: jshtsang@hku.hken_HK
dc.identifier.authorityTsang, JSH=rp00792en_HK
dc.description.naturepublished_or_final_versionen_HK
dc.identifier.doi10.1186/1471-2180-9-233en_HK
dc.identifier.pmid19878597-
dc.identifier.pmcidPMC2777183-
dc.identifier.scopuseid_2-s2.0-72849125655en_HK
dc.identifier.hkuros168290en_HK
dc.relation.referenceshttp://www.scopus.com/mlt/select.url?eid=2-s2.0-72849125655&selection=ref&src=s&origin=recordpageen_HK
dc.identifier.volume9en_HK
dc.identifier.eissn1471-2180-
dc.identifier.isiWOS:000272051200001-
dc.publisher.placeUnited Kingdomen_HK
dc.identifier.scopusauthoridTse, YM=35244048800en_HK
dc.identifier.scopusauthoridYu, M=7404272859en_HK
dc.identifier.scopusauthoridTsang, JSH=7102483508en_HK
dc.identifier.citeulike6186549-

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